1. Field of the Invention
The invention relates to a medicine, and in particular to azaazulene compounds which modulate the protein kinases (PKs) activity and/or treat cancer.
2. Description of the Related Art
Protein kinases (PKs) are enzymes which catalyze the phosphorylation of specific tyrosine, serine, or threonine residues in cellular proteins. The PKs mediate cellular signal transduction in regulating cellular function such as proliferation, differentiation, growth, cell cycle, cell metabolism, cell survival, cell apoptosis, DNA damage repair, cell motility, and response to the microenvironment. Disregulated PKs activity is a frequent cause of disease such as angiogenesis, cancer, tumor growth, tumor metastasis, atherosclerosis, age related macular degeneration, diabetic retinopathy, inflammatory diseases and/or parasitical disease.
The PKs can be divided into two classes: the protein tyrosine kinases (PTKs) and the serine/threonine kinases (STKs). PTKs, which catalyze the transfer of the gamma-phosphate of ATP to tyrosine residues in protein substrates is one of the key covalent modifications that occurs in multi-cellular organisms as a result of intercellular communication during embryogenesis and maintenance of adult tissues. Phosphorylation of tyrosine residues modulates enzymatic activity of PTKs and the recruitment of downstream signaling proteins. Two classes of PTKs are present in cells: the transmembrane receptor PTKs and the nonreceptor PTKs. PTKs are critical components of cellular signaling pathways, their catalytic activity is strictly regulated. Unregulated activation of these enzymes, through mechanisms such as point mutations or over-expression, can lead to various forms of cancer as well as benign proliferative conditions. The importance of PTKs in health and disease is further underscored by the existence of aberrations in PTK signaling occurring in inflammatory diseases and diabetes. The growth factor receptors with PTK activity are known as receptor tyrosine kinases (“RTKs”). They comprise a large family of transmembrane receptors with diverse biological activity. The intracellular kinase domains of RTKs can be divided into two classes: those containing a stretch of amino acids separating the kinase domain and those in which the kinase domain is continuous. Activation of the kinase is achieved by ligand binding to the extracellular domain, which induces dimerization or oligomerization of the receptors. Receptors thus activated are able to autophosphorylate tyrosine residues outside the catalytic domain via cross-phosphorylation. The results of this auto-phosphorylation are stabilization of the active receptor conformation and the creation of phosphotyrosine docking sites for proteins which transduce signals within the cell. Signaling proteins which bind to the intracellular domain of receptor tyrosine kinases in a phosphotyrosine-dependent manner include RasGAP, PI3-kinase, phospholipase C phosphotyrosine phosphatase SHP and adaptor proteins such as Shc, Grb2 and Crk.
The EGFR, epidermal growth factor receptor, belongs to a family of receptor tyrosine kinases in mammals which is composed of four members: EGFR (ErB1), ErB2, ErB3, and ErB4. EGFR is an 1186 amino acid residue transmembrane glycoprotein. It consists of an extracellular ligand binding domain, an intracellular tyrosine kinase domain, and a COOH terminal region that contains autophosphorylation sites. The binding of specific ligands, such as EGF, transforming growth factor-beta, betacellulin, heparin-binding EGF, epiregulin, or amphiregulin, results in phosphorylation of multiple tyrosine residues in the COOH-terminal tail, triggering the cellular signaling pathway that regulates fundamental cellular processes such as proliferation, migration, differentiation and survival. EGFR is over expressed in many types of tumor cells, such as bladder, lung, gastric, breast, brain, head & neck, cervix, ovary, endometrium, etc. Abnormally high EGFR activity can be characteristic of non-small-cell lung cancers, breast cancers, ovarian cancers, bladder cancers, prostate cancers, salivary gland cancers, pancreatic cancers, endometrial cancers, colorectal cancers, kidney cancers, head and neck cancers, and glioblastoma multiforme. A tyrosine kinase inhibitor targeted to EGFR can be used for the treatment of cancers having abnormally high EGFR kinase activity and EFGR kinase disorder diseases.
One of RTK subfamily is referred to as the platelet derived growth factor receptor (“PDGFR”) group, which includes PDGFR.alpha., PDGFR.beta., CSFIR, c-KIT and c-fms. These receptors consist of glycosylated extracellular domains composed of variable numbers of immunoglobin-like loops and an intracellular domain wherein the tyrosine kinase domain is interrupted by unrelated amino acid sequences. PDGFR signals induce expression of pro-angiogenic signals (including VEGF) in endothelial cells, further stimulating tumor angiogenesis. The PDGFR signaling pathway may play an important role in cell proliferation, cell migration, and angiogenesis, and may mediate the high interstitial fluid pressure of tumors.
Another group which, because of its similarity to the PDGFR subfamily, is sometimes subsumed into the later group is the fetus liver kinase (“flk”) receptor subfamily. This group is believed to be made up of kinase insert domain-receptor fetal liver kinase-1 (KDR/FLK-1, VEGF-R2), flk-1R, flk-4 and fms-like tyrosine kinase 1 (flt-1). Abnormally high PDGFR activity can be characteristic of gastrointestinal stromal tumor, small cell lung cancer, glioblastoma multiforme, and prostate cancer. A tyrosine kinase inhibitor targeted to PDGFR can be used for the treatment of cancers having abnormally high PDGFR kinase activity and PDGFR kinase disorder diseases.
FLT-3 (FMS-like tyrosine kinase 3) is a class III RTK structurally related to PDGFR, and colony stimulating factor 1 (CSF1). These RTK contain five immunoglobulin-like domains in the extracellular region and an intracellular tyrosine kinase domain split in two by a specific hydrophilic insertion (kinase insert). FLT-3 expression was described on bone marrow CD34-positive cells, corresponding to multipotential, myeloid and B-lymphoid progenitor cells, and on monocytic cells. FLT3 expression is restricted to cells of the fetal liver expressing high levels of CD34. FLT3 receptor function can be defined by the activity of its ligand (FL). FL is an early acting factor and supports the survival, proliferation and differentiation of primitive hemopoietic progenitor cells. Ligand binding to FLT3 promotes receptor dimerization and subsequent signaling through phosphorylation of multiple cytoplasmatic proteins, including SHC, SHP-2, SHIP, Cbl, Cbl-b, Gab1 and Gab2, as well as the activation of several downstream signalling pathways, such as the Ras/Raf/MAPK and PI3 kinase cascades. Internal tandem duplications (ITD) and/or insertions and, rarely, deletions in the FLT3-gene are implicated in 20-25% of all acute myeloid leukemias (AML). The duplicated sequence belongs to exon 11 but sometimes involves intron 11 and exon 12. The most frequently used nomenclature is FLT3-ITD. Because of the very heterogeneous molecular structure the term FLT3-LM (length mutation) seems to be more adequate. It was also described to be involved in 5-10% myelodysplastic syndromes (MDS) refractory anemia with excess of blasts (RAEB 1 and RAEB 2) and rare cases with acute lymphoblastic leukemia (ALL). A tyrosine kinase inhibitor targeted to FLT-3 can be used for the treatment of cancers having abnormally high FLT-3 kinase activity and FLT3 kinase disorder diseases.
C-KIT, SCFR (Stem Cell Factor Receptor), is known as type III receptor tyrosine kinase, structurally related to CSF-1R, PDGFR, and FLT-3, containing an extracellular domains with 5 Ig-like loops, a highly hydrophobic transmembrane domain, and an intracellular domain with tyrosine kinase activity split by a kinase insert (KI) in an ATP-binding region and in the phosphotransferase domain. C-Kit is expressed on the cell plasma membrane in the hematopoietic stem cells, mast cells, melanocytes, and germ-cell lineages. SCF/MGF receptor with PTK activity, binding of ligand (SCF) induces receptor dimerization, autophosphorylation and signal transduction via molecules containing SH2-domains. With the abnormal activity expression, mast cell hyperplasia in the bone marrow, liver, spleen, lymph nodes, gastrointestinal tract and skin, gain of function mutations are detected in most patients. It is recognized as clinical features of malignant hematopoietic cell growth are influenced by the time, the location of c-kit mutative events, and the number of associated lesions. A tyrosine kinase inhibitor targeted to c-Kit can be used for the treatment of cancers having abnormally high c-Kit kinase activity and c-Kit kinase disorder diseases.
Another member of the tyrosine kinase growth factor receptor family is the vascular endothelial growth factor receptor (VEGFR) subgroup. VEGFR is a dimeric glycoprotein similar to PDGFR but has different biological functions and target cell specificity in vivo. In particular, VEGFR is presently thought to play an essential role is vasculogenesis and angiogenesis. Angiogenesis is essential for tumor growth and survival. There are 3 distinct VEGF receptors-VEGFR-1, -2, and -3. Each of them contributes separately to the angiogenic process. VEGFR-1 is thought to play a role in regulating VEGF binding to VEGFR-2 during angiogenesis. VEGFR-2 (KDR) stimulates the proliferation, migration, and survival of endothelial cells during angiogenesis and is recognized as a critical VEGF receptor for angiogenesis. VEGFR-3 stimulates the proliferation, migration, and survival of endothelial cells during lymphangiogenesis, which in turn facilitates metastases. Despite these seemingly distinct roles, all VEGFRs overlap to some degree in their function, leading to significant redundancy. Therefore, inhibition of all identified VEGF receptors may ensure more complete inhibition of angiogenesis. A tyrosine kinase inhibitor targeted to VEGFR can be used for the treatment of solid tumors and vascular disorder diseases.
c-Met (hepatocyte growth factor receptor), is the high affinity receptor for HGF/SF, a multifunctional cytokine. Upon ligand binding, MET dimerizes and transphosphorylates tyrosine residues in the C-terminal domain, which then interacts with members of a variety of signaling pathways. These include Grb-2 associated binder 1, phosphoinositide 3′ kinase and c-Src. Under physiological conditions, MET-HGF/SF signaling has been shown to affect a wide range of biological activities depending on the cell target. These activities vary from cell proliferation (mitogenesis) to cellular shaping (morphogenesis) and motility (motogenesis). The coordination of these diverse activities constitutes a genetic program of invasive growth that allows branched morphogenesis (the formation of epithelial tubular structures), myoblast migration and neurite branching. MET/HGF cell targets comprise epithelial and mesenchymal cells, hematopoietic cells, myoblasts, spinal motor neurons. MET-HGF/SF signaling is also essential for normal development: mouse embryos carrying null mutations in both HGF alleles die in midgestation and show impaired liver formation. MET and its ligand hepatocyte growth factor/scatter factor (HGF/SF) are expressed in numerous tissues although predominantly in cells of epithelial and mesenchymal origin, respectively. MET is amplified and overexpressed in many types of tumors, including tumors of the kidney, thyroid, pancreas and osteosarcoma. A tyrosine kinase inhibitor targeted to c-Met can be used for the treatment of cancers having abnormally high c-Met kinase activity and c-Met kinase disorder diseases.
RET is a tyrosine kinase receptor whose ligands are neurotrophic factors of the glial-cell line derived neurotrophic factor (GDNF) family, including GDNF, neurturin, artemin and persefin. RET activation is mediated via different glycosyl phosphatidylinositol-linked GRF receptors. 3 main isoforms of RET is detected in human, such as long isoform (RET51): 1114 amino acids, middle isoform (RET 43): 1106 amino acids and short isoform (RET 9): 1072 amino acids. RET is mainly expressed in tumors of neural crest origin: medullary thyroid carcinoma, pheochromocytoma and neuroblastoma. In human embryos, RET is expressed in a cranial population of neural crest cells, and in the developing nervous and urogenital systems. RET expression is found in several crest-derived cell lines, spleen, thymus, lymph nodes, salivary glands, spermatogonia, and recently in normal thyroid tissue, thyroid adenoma and both papillary and follicular thyroid cell neoplasias. A tyrosine kinase inhibitor targeted to RET can be used for the treatment of cancers having abnormally high RET kinase activity and RET kinase disorder diseases.
c-ABL (v-abl Abelson murine leukemia viral oncogene homolog) exhibit a permanent nuclear and cytoplasmic shuttling activity, driven by 3 nuclear localization signals (NLS) and a single nuclear export signal (NES) close to the C-terminal region. BCR/ABL has a cytoplasmic localization role and all three BCR-ABL fusion proteins have been shown to exhibit oncogenic potential. All three hybrid proteins have increased protein kinase activity compared to ABL: 3BP1 (binding protein) binds normal ABL on SH3 domain, which prevents SHI activation. Nuclear and cytoplasmic ABL may have different functions. 1-Nuclear c-ABL plays a major role in the regulation of cell death after DNA damage. All DNA damage inducing agents activate nuclear c-ABL kinase in an ATM-dependent manner and in the presence of the p53-homolog p73 protein. The latter is physically associated with c-ABL after DNA damage through the SH3 domain of c-ABL. DNA damage also activates simultaneously p53 pathway, leading to the activation of Rb which induces growth arrest and protects cells from apoptosis. The exact mechanisms of apoptosis induced by c-ABL are unknown. The nuclear entrapment of BCR-ABL has also been shown to induce apoptosis in leukemic cells. 2-Cytoplasmic c-ABL: possible function in adhesion signaling as an efflux of c-ABL from nucleus to the cytoplasm is found in fibroblasts after adhesion. A tyrosine kinase inhibitor targeted to c-ABL can be used for the treatment of cancers having abnormally high c-ABL kinase activity and c-ABL kinase disorder diseases.
TIE (tyrosine kinase with immunoglobulin-like and EGF-like domains) can be defined into two subgroups. TIE-1 (tyrosine kinase with Ig and EGF homology domains 1) and TIE-2/Tek comprise a receptor tyrosine kinase (RTK) subfamily with unique structural characteristics: two immunoglobulin-like domains flanking three epidermal growth factor (EGF)-like domains and followed by three fibronectin type III-like repeats in the extracellular region and a split tyrosine kinase domain in the cytoplasmic region. These receptors are expressed primarily on endothelial and hematopoietic progenitor cells and play critical roles in angiogenesis, vasculogenesis and hematopoiesis. Human TIE-1 cDNA encodes a 1124 amino acid (aa) residue precursor protein with an 18 residue putative signal peptide, a 727 residue extracellular domain and a 354 residue cytoplasmic domain. Two ligands, angiopoietin-1 (Ang1) and angiopoietin-2 (Ang2), which bind TIE-1 with high affinity, have been identified. Ang2 has been reported to act as an antagonist for Ang1. A tyrosine kinase inhibitor targeted to TIE can be used for the treatment of solid tumors and vascular disorder diseases.
FGFR (fibroblast growth factor receptors) consist of an extracellular ligand domain comprised of three immunoglobulin-like domains, a single transmembrane helix domain, and an intracellular domain with tyrosine kinase activity. The fibroblast growth factors are the largest family of growth factor ligands comprising of 23 members. FGFRs share a similar sequence structure, characterized by three extracellular immunoglobulin-like domains (IgI, IgII, and IgIII), a single-pass transmembrane segment, and a split tyrosine kinase (TK1/TK2) domain. The great majority of pathogenic FGFR mutations are missense, and all confer gain of function to the mutated protein. Some mutations are highly recurrent. The gain-of-function mechanisms identified for FGFR2 mutations are (a) selectively enhanced FGF-binding affinity, (b) illegitimate FGF-binding specificity, (c) FGF-independent covalent dimerization, and (d) ectopic spliceoform expression. These mechanisms account for the dominant inheritance of all the associated phenotypes. A tyrosine kinase inhibitor targeted to FGFR can be used for the treatment of cancers having abnormally high FGFR kinase activity and FGFR kinase disorder diseases.
Insulin-like growth factor 1 (IGF1) was considered a potential candidate for the treatment of heart failure. However, some animal studies and clinical trials have questioned whether elevating IGF1 chronically is beneficial. Secondary effects of increased serum IGF1 levels on other tissues may explain these unfavorable results. The aim of the current study was to examine the role of IGF1 in cardiac myocytes in the absence of secondary effects, and to elucidate downstream signaling pathways and transcriptional regulatory effects of the IGF1 receptor (IGF1R). Activation of the IGF-1 receptor is survival and proliferation in mitosis-competent cells, and growth (hypertrophy) in tissues such as skeletal muscle and cardiac muscle. The IGFR signaling pathway is of critical importance during normal development of mammary gland tissue during pregnancy and lactation. Several growth factors and hormones are involved in this overall process, and IGF-1R is believed to have roles in the differentiation of the cells and a key role in inhibiting apoptosis until weaning is complete. The IGF-1R is implicated in several cancers, most notably breast cancer. It is further implicated in breast cancer by increasing the metastatic potential of the original tumor by inferring the ability to promote vascularisation. A tyrosine kinase inhibitor targeted to IGFR can be used for the treatment of cancers having abnormally high IGFR kinase activity and IGFR kinase disorder diseases.
Kinases such as c-Src, c-Abl, mitogen activated protein (MAP) kinase, phosphotidylinositol-3-kinase (PI3K) AKT, and the epidermal growth factor (EGF) receptor are commonly activated in cancer cells, and are known to contribute to tumorigenesis. Many of these occur in the same signaling pathway—for example, HER-kinase family members (HER1 [EGFR], HER3, and HER4) transmit signals through MAP kinase and PI3 kinase to promote cell proliferation.
TrkA (Tropomyosin-Related Kinase A) is a high affinity catalytic receptor for neurotrophin, Nerve Growth Factor (NGF) and thus mediates the multiple effects of NGF including neuronal differentiation and survival. The TrkA receptor is part of the large family of receptor tyrosine kinases.
PTK disorder disease includes, such as cancer, asarthritis, diabetic retinopathy, restenosis, hepatic cirrhosis, atherosclerosis, angiogensis, glomerulonephritis, diabetic nephropathy, thrombic microangiopathy syndromes, transplant rejection, autoimmune disease, diabetes, and hyperimmune disorders.
Cancers include, without limitation, carcinoma of the bladder, breast, colon, kidney, liver, lung, head and neck, gall-bladder, ovary, pancreas, stomach, cervix, thyroid, prostate, skin, hematopoietic tumor of lymphoid lineage (i.e. leukemia, acute lymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell-lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, hairy cell lymphoma and Burkett's lymphoma), hematopoietic tumor of myeloid lineage (i.e. acute myelogenous leukemia, chronic myelogenous leukemia, multiple myelogenous leukemia, myelodysplastic syndrome and promyelocytic leukemia), tumor of mesenchymal origin (i.e. fibrosarcoma and rhabdomyosarcoma), tumor of the central or peripheral nervous system (i.e. astrocytoma, neuroblastoma, glioma and schwannomas), melanoma, seminoma, teratocarcinoma, osteosarcoma, thyroid follicular cancer; and Kaposi's sarcoma.